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An Engineered Rare Codon Device for Optimization of Metabolic Pathways.

ABSTRACT: Rare codons generally arrest translation due to rarity of their cognate tRNAs. This property of rare codons can be utilized to regulate protein expression. In this study, a linear relationship was found between expression levels of genes and copy numbers of rare codons inserted within them. Based on this discovery, we constructed a molecular device in Escherichia coli using the rare codon AGG, its cognate tRNA (tRNA(Arg) (CCU)), modified tRNA(Asp) (GUC???CCU), and truncated aspartyl-tRNA synthetase (TDRS) to switch the expression of reporter genes on or off as well as to precisely regulate their expression to various intermediate levels. To underscore the applicability of our work, we used the rare codon device to alter the expression levels of four genes of the fatty acid synthesis II (FASII) pathway (i.e. fabZ, fabG, fabI, and tesA') in E. coli to optimize steady-state kinetics, which produced nearly two-fold increase in fatty acid yield. Thus, the proposed method has potential applications in regulating target protein expression at desired levels and optimizing metabolic pathways by precisely tuning in vivo molar ratio of relevant enzymes.

SUBMITTER: Wang Y

PROVIDER: S-EPMC4745014 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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An Engineered Rare Codon Device for Optimization of Metabolic Pathways.

Wang You Y Li Chunying C Khan Md Rezaul Islam MR Wang Yushu Y Ruan Yunfeng Y Zhao Bin B Zhang Bo B Ma Xiaopan X Zhang Kaisi K Zhao Xiwen X Ye Guanhao G Guo Xizhi X Feng Guoyin G He Lin L Ma Gang G

Scientific reports 20160208

Rare codons generally arrest translation due to rarity of their cognate tRNAs. This property of rare codons can be utilized to regulate protein expression. In this study, a linear relationship was found between expression levels of genes and copy numbers of rare codons inserted within them. Based on this discovery, we constructed a molecular device in Escherichia coli using the rare codon AGG, its cognate tRNA (tRNA(Arg) (CCU)), modified tRNA(Asp) (GUC → CCU), and truncated aspartyl-tRNA synthet ...[more]

PMID: 26852704

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Project description:BACKGROUND: During the last decade, a number of authors have shown that the genetic regulation of metabolic networks may follow optimality principles. Optimal control theory has been successfully used to compute optimal enzyme profiles considering simple metabolic pathways. However, applying this optimal control framework to more general networks (e.g. branched networks, or networks incorporating enzyme production dynamics) yields problems that are analytically intractable and/or numerically very challenging. Further, these previous studies have only considered a single-objective framework. RESULTS: In this work we consider a more general multi-objective formulation and we present solutions based on recent developments in global dynamic optimization techniques. We illustrate the performance and capabilities of these techniques considering two sets of problems. First, we consider a set of single-objective examples of increasing complexity taken from the recent literature. We analyze the multimodal character of the associated non linear optimization problems, and we also evaluate different global optimization approaches in terms of numerical robustness, efficiency and scalability. Second, we consider generalized multi-objective formulations for several examples, and we show how this framework results in more biologically meaningful results. CONCLUSIONS: The proposed strategy was used to solve a set of single-objective case studies related to unbranched and branched metabolic networks of different levels of complexity. All problems were successfully solved in reasonable computation times with our global dynamic optimization approach, reaching solutions which were comparable or better than those reported in previous literature. Further, we considered, for the first time, multi-objective formulations, illustrating how activation in metabolic pathways can be explained in terms of the best trade-offs between conflicting objectives. This new methodology can be applied to metabolic networks with arbitrary topologies, non-linear dynamics and constraints.

Dataset Information

An Engineered Rare Codon Device for Optimization of Metabolic Pathways.

Publications

An Engineered Rare Codon Device for Optimization of Metabolic Pathways.

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